WildC. elegansand other nematodes live in dirt and eat
bacteria, relying on mechanoreceptor neurons (MRNs) to detect collisions with soil
particles and other animals as well as forces generated by their own movement. MRNs may
also help animals detect bacterial food sources. Hermaphrodites and males have 22
putative MRNs; males have an additional 46 MRNs, most, if not all of which are needed
for mating. This chapter reviews key aspects ofC. elegansmechanosensation, including MRN anatomy, what is known about their contributions to
behavior as well as the neural circuits linking MRNs to movement. Emerging models of the
mechanisms used to convert mechanical energy into electrical signals are also discussed.
Prospects for future research include expanding our understanding of the molecular basis
of mechanotransduction and how activation of MRNs guides and modulates behavior.

1. Introduction

Wild C. elegans and other free-living nematodes live in dirt and
eat bacteria. In this complex, three-dimensional environment, worms experience external
forces produced by surface tension, by colliding with adjacent soil particles and other
animals, as well as forces generated by their own movement. Hermaphrodites have 30
sensory neurons that might be recruited to detect these forces, while males have 52
additional putative mechanoreceptor neurons (MRNs; Table 1).

Table 1. Putative C. elegans mechanoreceptor neurons (MRNs).

Hermaphrodite MRNs (30)

Male-specific MRNs (52)

body wall

Touch receptor neurons:

2 ALM, 2 PLM, AVM, PVM

2 PVD

2 ADE

2 PDE

fan

Ray Neurons:

18 RnA, 18 RnB

hook

1 HOA

1 HOB

nose tip

2 ASH

2 FLP

4 OLQ

4 CEP

6 IL1

spicule

2 SPD

2 SPV

2 PCA

2 PCC

4 SPC*

*SPC neurons innervate the protractor muscles in males and have been proposed to act as proprioceptors because they are attached
to the protractor and anal depressor muscles (Sulston et al., 1980).

Several laboratory assays of behavioral responses to touch have been developed for C.
elegans see the Behavior section in WormMethods. These assays, especially the assay for responses
to gentle touch, have provided critical tools for the study of mechanosensation. As a
result, more is known about the molecules needed for the function, specification, and
development of C. elegans MRNs than is known in other organisms.
This chapter reviews key aspects of MRN function, focusing on the anatomy of MRNs,
molecules proposed to mediate the conversion of mechanical energy into electrical
signals, and the neural networks linking activation of MRNs to behavior. To learn more
about similarities with mechanosensation in other organisms, the reader should consult
the following recent review articles: Ernstrom and Chalfie, 2002;
Goodman and Schwarz, 2003; Syntichaki and Tavernarakis, 2004.

2.
C. elegans mechanoreceptor neurons

In one sense, all mechanoreceptor neurons (MRNs) are similar: they generate electrical
signals in response to mechanical energy and transmit those signals to other neurons via
electrical and/or chemical synapses. Yet, in another sense, they differ. Some are
specialized to detect external mechanical stimuli, while others inform the nervous
system about self-generated stimuli (e.g., stretch receptors). The sensory dendrites of
MRNs may be ciliated or nonciliated and, if ciliated, they can be encapsulated within
the cuticle or exposed to the external environment. Each neuron is located in a specific
place in the body, optimized to detect forces delivered to that part of the body.

2.1. Nonciliated MRNs

2.1.1. Touch receptor neurons

2.1.1.1. What do touch receptor neurons look like?

The six touch receptor neurons (ALML, ALMR, AVM, PLML, PLMR, PVM) extend
long processes that innervate approximately one-half of the animal’s body
length (Figure 1). Their processes are filled with unusual,
15-protofilament (15-p) microtubules that are cross-linked to one another
and arrayed such that their distal ends are in close apposition to the cell
membrane (Chalfie and Thomson, 1979). The 15-p microtubules are
unique to the touch receptor neurons (Chalfie and Sulston, 1981) and arise from tubulins (MEC-7 and MEC-12) abundant in touch
receptor neurons (Fukushige et al., 1999; Savage et al., 1989). Ultrastructural specializations are not restricted to the
cytoskeleton, however. Touch receptor neurons are tightly coupled to the
animal’s skin or cuticle, engulfed by hypodermal cells, and surrounded by an
electron-dense extracellular matrix or ECM (Chalfie and Sulston, 1981). Because they are the only neurons whose processes are in such
close contact with the cuticle along their length, touch receptor neurons
are uniquely positioned to detect external forces applied to the animal’s
body wall, as well as internal forces generated during locomotion.

Figure 1. Gross and fine morphology of the touch receptor neurons. Position of the cell bodies and neuronal processes of the touch receptor neurons, only
the left side of the animal is shown (left) and electron micrograph of a PLM neuron
(right). The electron micrograph is from an L4 animal, courtesy of J. Cueva, Stanford
University.

2.1.1.2. What behaviors do they mediate?

Intact touch receptor neurons mediate behavioral responses to touches
delivered by an eyebrow hair to the body wall. Such responses are described
as simple avoidance behaviors and defined by a differential assay (see the Behavior section in WormMethods). Touch a worm anywhere along the anterior half of the body wall
and it will back away (Movie 1); touch a backward moving
worm along the posterior half of the body wall and it will move forward. If
the animal is already moving forward, it will accelerate in response to a
posterior touch. Animals that fail to respond to stimuli delivered by an
eyebrow hair (gentle touch) retain sensitivity to stimuli delivered by
prodding with a wire pick (harsh touch). Responses to anterior touch require
ALML, AMLR, and AVM, while responses to posterior touch require PLML and
PLMR (Chalfie et al., 1985; Wicks et al., 1996).
A role for PVM in touch withdrawal has not been demonstrated (Chalfie et
al., 1985; Wicks et al., 1996). Substrate
vibrations produced by plate tap are believed to activate all touch receptor
neurons simultaneously. Thus, behavioral responses to plate tap reflect a
combination of the antagonistic reflexes, one initiated by anterior touch
and activation of ALML/R and AVM and a second one initiated by posterior
touch and activation of PLML/R. Stimulation of the anterior touch receptor
neurons (ALML/R and AVM) dominates in adults— i.e. wild-type adults reverse
direction in response to plate tap (Wicks et al., 1996).

Worms habituate to repeated plate tap, a process that requires intact
touch receptor neurons (Wicks et al., 1996). The shorter the
inter-stimulus interval, the faster habituation occurs (Rankin and Broster,
1992). Mutations that disrupt signaling by glutaminergic and
dopaminergic neurons (eat-4 and
cat-2, respectively) accelerate short-term habituation
without affecting responses to single tap stimuli (Rankin and Wicks, 2000; Sanyal et al., 2004). These observations
implicate the circuit linking touch to locomotion in the regulation of
short-term habituation. The effect of cat-2 (which
encodes tyrosine hydroxylase, an enzyme required for dopamine biosynthesis)
is particularly intriguing and is proposed to involve neurohumoral effects
of dopamine on the touch receptor neurons (Sanyal et al., 2004). Short-term behavioral habituation might also reflect
de-sensitization of the touch receptor neurons. Indeed, trains of 30 stimuli
are sufficient to decrease touch-evoked changes in intracellular
Ca2+ (Suzuki et al., 2003).
Mechanoreceptor current amplitude does not decline in response to trains of
up to 60 stimuli, however (O'Hagan et al., 2005). Considered
together, these results suggest that short-term behavioral habituation
reflects plasticity in signaling processes subsequent to transduction in the
touch receptor neurons and in the circuit linking them to locomotion.

Movie 1. This movie shows the response of a wild-type (N2) worm to mechanical stimuli
delivered to the head. In this movie, a calibrated microforce probe delivering ~10
μN of force was used.

The PVD cells have long anterior and posterior neurites that branch to
form a complex dendritic network blanketing the body surface (Halevi et al.,
2002; Tsalik et al., 2003). Since the PVD
neurons were not completely reconstructed by White et al. (White et al.,
1986), this branching pattern was not appreciated until recently.
It is reminiscent of multi-dendritic sensory neurons that innervate the
surface of insect larvae (Bodmer and Jan, 1987) and is consistent with
the idea that PVD is a MRN. In addition to this dendritic network, PVD
neurons have a ventrally directed neurite that synapses onto interneurons
that control locomotion.

Strong mechanical stimuli (e.g., prodding the body wall with a wire)
produce an avoidance response that persists in animals that lack touch
receptor neurons. Killing the PVD neurons in such animals disrupts this
behavior, suggesting that PVD neurons are high-threshold MRNs. Mutations in
the mec-3 gene, which encodes a LIM domain
transcription factor expressed in the touch receptor neurons and PVD,
eliminate responses to both weak and strong mechanical stimuli (Way and
Chalfie, 1989). mec-3 animals are
insensitive to weak stimuli because their touch receptor neurons fail to
differentiate properly. They may be insensitive to strong stimuli because
mec-3 appears to disrupt PVD’s dendritic network
(Tsalik et al., 2003). Mutations in the glr-1
ionotropic glutamate receptor also disrupt responses to strong
stimuli (Hart et al., 1995), probably because loss of
glr-1 affects signaling from PVD to interneurons
regulating locomotion.

The extensive PVD dendritic network suggests that it could be activated by
stimuli delivered anywhere along the body wall. PVD appears to detect forces
greater than ~100 μN. This estimate was obtained by comparing the touch
sensitivity of wild-type worms, which respond to 10 μN touches, with worms
lacking function touch receptor neurons, which fail to respond to forces
less than ~100 μN (Iris Chin, M.B.G., and Marty Chalfie, unpublished).

2.2. Ciliated MRNs

2.2.1. ASH

2.2.1.1. What does ASH look like?

The ciliated endings of the ASH neurons are located
in the left and right amphids (Figure 2), where they are
exposed to the external environment through a channel in the amphid sheath cell
(Ward et al., 1975; White et al., 1986). ASH neurons
accumulate fluorescent dyes via their exposed ciliated endings and screens for
defects in dye-filling have been used to identify genes needed for cilia
formation (Perkins et al., 1986).

ASH was classified as a primary sensory neuron based on its morphology and
identified as an MRN by laser ablation analysis. Support for this idea comes
from experiments that recorded electrical responses to nose touch in AVA, an
interneuron that receives direct synaptic input from ASH (Mellem et al.,
2002). Additional support comes from in
vivoCa2+ imaging (Hilliard et al.,
2005). Using cameleon, a genetically-encoded
Ca2+ indicator (Miyawaki et al., 1997; Nagai et al., 2004), Hilliard et al. (Hilliard et
al., 2005) found that nose touch produces
Ca2+ transients in ASH cell bodies, but only in
the presence of exogenous serotonin (5-HT). Assuming that sensitivity to
nose touch is food dependent, this finding is explained by a model in which
5-HT signals the presence of food and that exogenous 5-HT is an effective
substitute for food (Chao et al., 2004).

The four CEP neurons have ciliated dendrites distributed around the mouth,
embedded in sensilla next to those that contain the OLQ endings (Figure 2). By contrast, the ADE and PDE neurons have ciliated
dendrites embedded in the cuticle along the left and right lateral midlines.
These putative MRNs are the only dopaminergic neurons in the hermaphrodite
nervous system (Sulston et al., 1975).

2.2.3. Male-specific MRNs

2.2.3.1. What do they look like?

Forty-two male-specific MRNs are ciliated sensory neurons that innervate
the male tail, hook, post-cloacal sensilla, and spicules (Figure 3). The hook sensilla contain two sensory neurons (HOA
and HOB), while the spicule is innervated by four neurons (SPDL, SPDR, SPVL,
SPVR). Thirty-six male-specific sensory neurons innervate nine bilateral
pairs of sensory rays in the male tail (the RnA and RnB neurons, where ‘n’
indicates the ray number). They are exposed to the external environment in
channels, except for the neurons that innervate ray 6 (which are
encapsulated). Rays 1, 5, and 7 open to the dorsal side, while rays 2, 4,
and 8 open ventrally. Despite looking alike, the ray sensory neurons differ
with respect to their neurotransmitter phenotype. RnA in rays 5, 7, and 9
are dopaminergic, while RnB in rays 1, 3 and 9 are serotinergic. In some
rays, the amine transmitters are co-expressed with the
flp genes encoding FMRFamide-like peptides (Lints et
al., 2004).

Figure 3. Sensory rays in the male tale. a. lateral view. Blue indicates rays that open to the dorsal side, red indicates rays
that open to the ventral side, ray 6 is encapsulated and labeled in green. Amine
neurotransmitters indicated below. b. ventral view. Adapted from Goodman and Schwarz
(2003), with permission.

2.2.3.2. The role of MRNs in male mating behavior

Male C. elegans execute a series of stereotyped
behaviors during mating. Putative MRNs in the male tail play essential roles
in each step of mating behavior, as determined by observing the effects of
laser ablation on mating. The sensilla that innervate the ventral surface of
the male tail are required for responses to ventral contact with
hermaphrodites, while dorsally-directed rays mediate responses to dorsal
contact with hermaphrodites (Liu and Sternberg, 1995). The
hook sensilla function in vulva location, while the sensory neurons that
innervate the spicule are likely to provide feedback for spicule insertion
into the vulva and subsequent sperm release. It will be interesting to learn
more about how the sensory information provided by the male tail is
integrated and used to coordinate mating behavior.

3. Neural circuits linking mechanosensation to locomotion

The neural circuits linking mechanosensation to locomotion in C.
elegans hermaphrodites have been deduced by integrating the effects of
killing individual neurons on touch-mediated behaviors (Chalfie et al., 1985; Kaplan and Horvitz, 1993; Wicks and Rankin, 1995)
with the wiring diagram described in the ‘Mind of the Worm’ (White et al., 1986; Figure 4). A common thread are the four pairs of command
interneurons: AVA, AVB, AVD, and PVC. AVA and AVD are needed for backward locomotion,
while AVB and PVC appear to mediate forward movement.

Mechanical stimuli delivered to the nose and head elicit reversals that are frequently
terminated by ‘omega turns’ (Croll, 1975). How activation of anterior MRNs
leads to omega turns is unclear, however. One possibility is that MRNs signal to the SMD
and RIV motor neurons, recently demonstrated to be important for omega turns (Gray et
al., 2005). For stimuli that activate either the ASH neurons or the
anterior touch receptor neurons, such signaling is likely to be polysynaptic, since
there are no direct connections between either class of MRN and the SMD and RIV motor
neurons (White et al., 1986).

4. Molecules and mechanisms of mechanotransduction

Genetic analysis of touch sensation in worms and fruit flies has revealed subunits of
putative transduction complexes (Ernstrom and Chalfie, 2002; Goodman and
Schwarz, 2003). Most, if not all, proteins proposed to form transduction
channels in ciliated MRNs are members of the TRP channel superfamily. TRP proteins are
named for the trp (transient receptor potential) gene in
Drosphila. A total of 24 C. elegans genes
are predicted to encode TRP channels (Table 2: Goodman and Schwarz, 2003). This prediction has yet to be confirmed by showing that C.
elegans TRPs form ion channels in heterologous cells, however. Candidate
transduction channels that operate in nonciliated MRNs include members of the DEG/ENaC
superfamily. Twenty-eight C. elegans genes encode members of this
superfamily (Table 2; Goodman and Schwarz, 2003), named for founding
members in C. elegans that can mutate to cause cellular
degeneration (‘degenerins’) and vertebrate epithelial Na channels (‘ENaCs’). Three
C. elegans DEG/ENaCs have been shown to form homomeric or
heteromeric amiloride-sensitive Na+ channels in heterologous cells: UNC-105, MEC-4, and
MEC-10 (Garcia-Anoveros et al., 1998; Goodman et al., 2002).

Additional TRP channels that might contribute to mechanosensation include a
C. elegans TRPN and TRPP protein. In
Drosphila and zebrafish, TRPN1 (called NOMPC in
Drosphila) is required for normal electrical responses to
mechanical stimuli in mechanosensory epithelia (Sidi et al., 2003; Walker
et al., 2000). trp-4 (also called CeNOMPC) encodes a
TRPN protein expressed in the CEP and ADE mechanoreceptor neurons (Walker et al., 2000). No analyses of its contribution to the function of CEP and ADE in
texture sensing have been reported, however. pkd-2 encodes a TRPP
protein needed for male mating response and vulva location (the Lov phenotype) expressed
in two classes of MRNs in the male tail (Barr et al., 2001): the ray
sensory neurons and HOB neurons. PKD-2 is co-expressed with LOV-1 and, together, these
proteins may form a sensory mechanotransduction channel in male-specific MRNs. The human
orthologs of LOV-1 and PKD-2 appear to have this function in the primary cilium of
kidney epithelial cells (Nauli et al., 2003).

MEC-4, MEC-1 (fused to GFP), and MEC-5 (fused to CFP) puncta are aligned with
cuticular annuli and with intermediate filaments [visualized with MH4 staining, (Francis
and Waterston, 1991)] that mark hemidesmosome-like attachments in the
hypodermis (Emtage et al., 2004). This observation raises the possibility
that complexes that connect touch receptor neurons to the cuticle have a dual role in
attachment and mechanotransduction.

Other C. elegans DEG/ENaC proteins implicated in mechanosensation
include UNC-105 and UNC-8 which are expressed in body wall muscle and motor neurons,
respectively (Garcia-Anoveros et al., 1998; Liu et al., 1996; Tavernarakis et al., 1997). Both channels have been proposed to
act as stretch receptors—i.e. the channels are predicted to be activated by cell
stretch. While initial attempts to observe such currents in wild-type muscle cells have
been unsuccessful, amiloride-sensitive Na+ currents have been observed in
gain-of-function unc-105(n506) mutant muscle cells (Jospin et al., 2004).
Double mutant unc-105(n506); let-2(n281) muscle cells lack this
current, consistent with the previous observation that let-2
supresses the hypercontraction phenotype of unc-10(n506) (Liu et
al., 1996). No measurements of either native or expressed UNC-8-containing
channels have been reported. Evidence that UNC-8 might participate in mechanosensation
by motor neurons comes primarily from the observation of locomotion defects in
unc-8 null mutants (Tavernarakis et al., 1997).

5. Conclusions

Much has been learned, but still more remains to be discovered about mechanosensory
signaling and how such signals guide behavior. C. elegans offers an
unusual opportunity to take an integrated approach to this task: existing tools permit
us to study mechanosensation by observing how behavior is affected by touch, by
measuring cellular responses to mechanical stimuli using in vivoCa2+ imaging and patch-clamp recording, by searching for
molecules needed for MRN function, and by manipulating cell function using cell-specific
expression of engineered and heterologous proteins. Genetic screens will recover only a
fraction of the genes needed for MRN function. For example, screens will miss genes with
redundant or pleitropic functions. Cell-specific gene expression profiling is an
alternative approach that circumvents these complications. Several new genes likely to
be needed for the function of touch receptor neurons were uncovered using this approach
(Zhang et al., 2002). Challenges for the future include building complete
lists of the genes needed for the specification, development and function of each MRN
and improved understanding of how mechanosensation affects behavior. Additional
challenges involve understanding exactly how mechanical energy opens ion channels, the
role of ECM components in force transfer, and delineating differences in the mechanism
of mechanotransduction between ciliated and nonciliated MRNs.

6. Acknowledgements

I thank Erich Schwarz and two anonymous reviewers for comments; Monica Driscoll and
Josh Kaplan for the sketch that inspired Figure 4. Research in my laboratory is
supported by funding from the Alfred P. Sloan Foundation, the Donald B. and Delia E.
Baxter Foundation, and NINDS (RO1 NS047715-01).

Sawin, E.R., Ranganathan, R., and Horvitz, H.R. (2000). C. elegans locomotory rate is modulated by the environment through a dopaminergic pathway and by experience through a serotonergic pathway.
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